Metastatic castration resistant prostate cancer (mCRPC) remains incurable. Once patients have progressed beyond enzalutamide and abiraterone, there are limited options other than docetaxel. On the other hand, mCRPC is an appropriate target for radioimmunotherapy (RIT), especially with short-range, high linear energy transfer alpha-emitters (e.g., 212Pb), based on the pattern of small spreads, including bone marrow and lymph nodes, sites that typically receive high levels of circulating antibody. Given the demonstrated safety and efficacy of recently approved alpha-particle 223Ra for treatment of bone metastases in prostate cancer, alpha-particle radiotherapy combined with antibody specifically targeting to tumor cells hold great promise for therapy development against mCRPC. Antibodies that are rapidly internalized into tumor cells have improved tumor targeting efficiency. We have identified a panel of single chain antibodies (scFvs) selected from phage display libraries against human prostate cancer tissues. In vivo studies of one antibody labeled with 99mTc have shown excellent tumor specific targeting with high tumor uptake and tumor/non- target ratio in preclinical models of mCRPC. We have since identified the target antigen as CD46, a negative regulator of the innate immunity. Analysis of mCRPC clinical data shows that CD46 is homogeneously overexpressed by mCRPC and further upregulated in patients who are resistant to enzalutamide and abiraterone. CD46 is an exciting new cell surface antigen for therapy development against mCRPC. We hypothesize that optimized antibody formats, derived from our novel anti-CD46 internalizing human scFv, will improve tumor targeting in vivo; and when labeled with the emerging alpha-particle isotope (212Pb), CD46 antibody will be developed into a novel RIT for mCRPC. The goal of this project is to develop a series of novel antibody formats composed of antibody fragments and full-length antibodies, labeled them with alpha emitters, identified the optimized format for RIT. Through in vitro evaluation of tumor cell- specific binding and internalization and in vivo determination of tumor targeting and radiation dosimetry, the lead antibody will be evaluated for efficacy and toxicity in both localized and disseminated/metastatic mCRPC cell line xenograft models, and further in mCRPC patient-derived xenograft models. If successfully carried out, our study will identify a lead RIT agent for therapeutic development against mCRPC, addressing a dire need for patients who are either non-responsive or have developed resistant to enzalutamide and abiraterone.